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At least 865 records · Page 48Linked to original sources
PubMedDate not supplied

[Determination of the rate constants for the interaction of enzymes with substrates and inhibitors during a short incubation time].

Kinetics of enzymatic reactions was studied using equipment which permitted to decrease the time of incubation of an enzyme with substrate or inhibitor up to two sec; principles of action, design and experimental procedure are described. A method of pipetting under pressure was employed to accelerate an addition of suitable reagents to the enzyme solution and for rapid stirring of the reaction mixture. Time of incubation was automatically measured with a precision of plus or minus 0.01 sec using an electronic stop-watch. Start and stop of the watch were made by means of electric gauges with platinum electrodes, placed into the channels through which the substrate or inhibitor solutions were supplied to the reaction mixture.

Cholinesterase Inhibitors↗
PubMed2001

Automation and high through-put for a DNA database laboratory: development of a laboratory information management system.

Automation and high through-put production of DNA profiles has become a necessity in every DNA database unit. In our laboratory we developed a Laboratory Information Management System (LIMS) controlled workflow architecture, which comprises a robotic DNA extraction- and pipetting-system and a capillary electrophoresis unit. This allows a through-put of 4,000 samples per person per year. Improved sample handling and data management, full sample- and batch-histories, and software-aided supervision of result data, with a consequent average turn-around time of 8 days, are the main features of our new system.

Austria↗
PubMed2001

Activation of cumulus-free equine oocytes: effect of maturation medium, calcium ionophore concentration and duration of cycloheximide exposure.

Two different culture media (TCM-199 and follicular fluid), two activation treatments (10 and 50 micromol calcium ionophore l(-1)) and three culture periods with cycloheximide were evaluated to find effective culture conditions for activation of cumulus-free equine oocytes. Oocytes were collected by scraping the follicle walls of ovaries obtained from an abattoir. Oocytes with expanded cumuli were matured at 38.2 degrees C in a humidified atmosphere of 5% CO(2) in air, in either TCM-199 with 10% fetal bovine serum (FBS) and 5 microU FSH ml(-1), or in 100% follicular fluid derived from a preovulatory follicle 24 h after injection of hCG. After 40--42 h of in vitro maturation, oocytes were denuded by gentle pipetting in TCM-199 plus 10% FBS with hyaluronidase. Oocytes with intact cytoplasmic membranes (n = 398; 94% presumed metaphase II) were treated in protein-free PBS with 10 or 50 micromol calcium ionophore l(-1) for 5 min. After washing, the oocytes were cultured in TCM-199 containing 10% FBS and 10 microg cycloheximide ml(-1) for 6 h, in cycloheximide for 6 h and then in cycloheximide-free medium for 18 h, or in cycloheximide for 24 h. The oocytes were fixed and evaluated by fluorescence microscopy. Oocytes with pronucleus I--II (dense to decondensing chromatin), pronucleus III--IV (decondensed chromatin) or progressing towards the first cleavage division were considered activated. The activation rate for oocytes matured in TCM-199 was significantly (P < 0.05) higher than for oocytes matured in follicular fluid (49% (99/204) versus 35% (60/171), respectively; P < 0.05). Culture with cycloheximide for 24 h resulted in a significantly higher rate of activation (67%, 74/111) than did the 6 h (33%, 44/136) or 6 h plus 18 h (32%, 41/128) treatments. The highest rate of activation (82%) was observed in oocytes matured in TCM-199, treated with 50 micromol calcium ionophore l(-1) and cultured with cycloheximide for 24 h.

Animals↗
PubMed1975

Mass reconstitution of neutral cephalothin sodium using tandem sequence filling.

A procedure is described for reconstituting and packaging neutral cephalothin sodium in large numbers of underfilled minibags for i.v. bolus administration. A special rack to hold six 20-g Keflin vials was designed for the procedure which employs an automatic pipetting device in a laminar air flow hood. A reconstitution deck and needle holder are used in the admixing stage. The minibags are frozen and later thawed prior to dispensing. The average labor and material cost (excluding drug and minibag) ranges from $0.195 to $0.124 per minibag depending on batch size. The 20-g neutral cephalothin sodium vial improves speed of reconstitution and does not require transfer into an intermediate container during the packaging cycle.

Cephalothin↗
PubMedDate not supplied

Determination of selenium in marine biological tissues by transverse heated electrothermal atomic absorption spectrometry with longitudinal Zeeman background correction and automated ultrasonic slurry sampling.

A fast, sensitive, and reliable method for determination of selenium in marine biological tissues by electrothermal atomic absorption spectrometry with slurry sampling was developed. Slurries were prepared from fresh and frozen seafood samples that were previously homogenized, dried, and ground; particle sizes <100 microm were taken for analysis. A 3% (v/v) HNO3 solution containing 0.01% (v/v) Triton X-100 was used as slurry diluent. Slurries were mixed on an automated ultrasonic slurry sampler at 20% amplitude for 30 s just before an aliquot was injected into the furnace. The method was successfully validated against the following certified reference materials: NRCC CRM DORM-2 (Dogfish muscle); NRCC CRM TORT-2 (Lobster hepatopancreas); NRCC CRM DOLT-2 (Dogfish liver); and BCR CRM 278 (Mussel tissue), and was subsequently applied to determination of Se in 10 marine biological samples. The influences of the drying procedure (oven-, microwave-, and freeze-drying), matrix modifier amount, mass of solid material in cup, and pipetting sequence are discussed. The limit of determination of Se was 0.16 microg/g and the repeatability, estimated as between-batch precision, was in the range of 4-8%. Se contents in the samples ranged from 0.6 to 2.8 microg/g. The proposed method should be useful for fast assessment of the daily dietary intake of Se.

Animals↗
PubMed1975

[Poultry vaccination against N.C.D. by means of an intramuscular injection with LaSota strain vaccine; a practical investigation (author's transl)].

An intramuscular N.D.C. vaccination with LaSota vaccine, in animals of less than ten weeks which were previously vaccinated by the spray method at about 5 days and 25 days against N.C.D., leads to an immunity of at least three months, as expressed by a high concentration of haemagglutination inhibiting antibodies in the circulatory system. If the vaccinated animals are older than twenty weeks, the resulting immunity is very high indeed and shows practically no impairment within a year after vaccination (longer periods have not been investigated), so that it may be assumed that revaccination will not be necessary during the laying period. Antibody variations between different animals of a flock are slight. The residual immunity of previous vaccinations did not appear to affect the development or maintenance of the antibodies. No spread of vaccine was observed. Here it should be noted that animals with a low residual immunity were used for control. It would be desirable to use specifically pathogenfree animals for this purpose, but this is not really practicable in field conditions. Half a hen dose of the LaSota vaccine, dissolved in 1/2 ml physiological saline, was injected into the pectoralis muscle with the help of the Cornwall Pipetting Unit. Twenty flocks, involving 6900 animals, were vaccinated in field conditions. Not a single vaccination gave rise to any vaccination reaction. The method does have a high labour content; which is an objection.

Animals↗
PubMed1975

Enzymatic immunoassay vs. gas/liquid chromatography for determination of phenobarbital and diphenylhydantoin in serum.

The antiepileptic drugs diphenylhydantoin and phenobarbital were measured in serum by a new commercially available enzyme immunoassay procedure ("EMIT," Syva Corp.). The procedure requires less than 5 min and no more than 50 mul of serum per determination. It is simple; only four steps (pipetting and diluting with an automatic pipettor-dilutor) are required before spectrophotometry. Twenty replicate analyses of a serum containing phenobarbital and diphenylhydantoin gave results with a CV of 6.8% and 9.1%, respectively. Results attained in a large series of patients were compared with results by a gas/liquid chromatographic procedure. For phenobarbital r = 0.97, and for diphenylhydantoin r = 0.98. No false negatives or false positives were encountered.

Chromatography, Gas↗
PubMed1975

[Comparative studies on 2 radioimmunoassay technics for the rapid determination of human chorionic somatomammotropin (HCS) in normal early pregnancies].

To measure HCS concentrations in serum in early pregnancies, 2 commercially available kits (Pharmacia, Frankfurt, and Amersham-Buchler, Braunschweig) were used. When comparing them, we found that they were nearly of the same quality and equally suited for the measurement of HCS in early pregnancy. We ourselves should prefer to use the Amersham-Buchler kit, because it uses 2 standard curves, covering 2 ranges and thus avoiding to tilute the patients' sera, which decreases the possible pipetting errors and increases the accuracy of the determinations.

Female↗
PubMedDate not supplied

[Screening and detection of amphetamine derivatives in biological stains].

A GC/MS-method is described for the screening, detection and determination of the commonly used drugs amphetamine, methamphetamine, MDA, MDMA and MBDB in small blood samples and bloodstains using solid phase SPE columns and a pipetting robot (Gilson Aspec XL). The detection limits are in the order of 0.03 to 0.08 microgram/L and the correlation factors between 0.9982 and 0.9998. Furthermore the stability was investigated covering a storage time of 64 days. The method has proven useful in forensic cases with only small sample volumes or bloodstains.

3,4-Methylenedioxyamphetamine↗
PubMed1976

Measurement of diphenylhydantoin in 0.1-ml plasma samples: gas chromatography and radioimmunoassay compared.

Concentrations of diphenylhydantoin in 364 plasma samples have been measured both by radioimmunoassay and with a recently developed gas-chromatographic method, which requires only 0.1 ml of plasma per determination. There was an excellent correlation between values obtained by the two methods (r = 0.986), and in only 11 plasma samples did the results differ by more than 20%. Of the investigated samples, 105 were obtained from uremic patients. For these, an equally good agreement was obtained between the two methods. Within-assay variance was 3.1% for the immunoassay and 3.3% for the gas-chromatographic procedure. Without automatic pipetting equipment, the radioimmunoassay procedure took twice as long as the chromatographic assay, and the cost of chemicals was considerably higher. Nevertheless, the better sensitivity of the radioimmunoassay makes it of great value, especially in children, because plasma samples of 10 to 20 mul can be used.

Child↗
PubMed2002

Correlation of blood volume values and pulmonary artery catheter measurements.

OBJECTIVE: To correlate the values generated by direct total blood volume measurement with pulmonary artery catheter parameters and commonly used laboratory values in the management of critically ill patients. METHODS: This study was carried out at the Lutheran Medical Center, Brooklyn, New York, United States of America, during the period 1998-1999. We prospectively correlated the total blood volume (TBV) values generated by the blood volume analyzer (BVA)-100 using I131-tagged albumin, with the values obtained from pulmonary artery catheter (PAC) of central venous pressure, pulmonary capillary wedge pressure, cardiac output, and with laboratory values of hematocrit, lactate, arterial blood gas and mixed venous blood, in critically ill patients. Twenty-four intensive care unit (ICU) patients were studied. INCLUSION CRITERIA: Admission to the intensive care, pulmonary artery catheter insertion and (APACHE) II Acute Physiology and Chronic Health Evaluation score of 8-30 (mean=17.875). EXCLUSION CRITERIA: Pediatric patients, hemodynamically normal or stable patients, pregnancy, and critically ill patients that were managed in an ICU setting without PAC catheter. Height and weight were recorded. After the collection of an initial blood sample (5 cc), one cc of I131-tagged albumin (15-25 microcuries) was injected using a patented syringe. Five venous samples were collected after the isotope injection. RESULTS: The collection times were entered into the BVA-100. Hematocrit measurements were performed in duplicate. Blood samples were centrifuged and one ml from the plasma of each sample was pipetted (in duplicate) into the sample tube then placed into the BVA-100. The results showed that the TBV did not correlate with either pulmonary capillary wedge pressure or central venous pressure, and except for the cardiac output, there is no correlation between pulmonary capillary wedge pressure readings or TBV results and the other parameters considered in this study. CONCLUSION: This method can be released from the research fields and can be safely incorporated into the clinical arena. It provides an accurate assessment of the volume status in intensive care unit patients.

Adult↗
PubMed2000

[Determination of plasma homocysteine by high performance liquid chromatography with fluorescence detection].

This article report a highly sensitive method specific for the determination of homocysteine in plasma by high performance liquid chromatography with fluorescence detection. Half mL plasma with 100 microL 0.11 mol/L sodium borohydride in 50 mmol/L Tris-HCl (pH 9.0) was kept at 30 degrees C for 30 min, and then 0.5 mL 0.5 mol/L perchloric acid was added. After the mixture was kept at room temperature for 10 min and centrifuged at 15,000 r/min for 10 min, 0.5 mL aliquots of the supernatant solution was pipetted into another vial containing 0.1 mL 3 mmol/L Bromobimane in 1.0 mol/L Na2EDTA (pH 7.0) and 0.7 mL of 90 mmol/L ammonium bicarbonate buffer containing 1.43 mol/L Na2EDTA, pH 8.0. The content was mixed, kept at 37 degrees C for 30 min and centrifuged at 15,000 r/min for 10 min. Then 10 microL aliquots of the supernatant solution was injected into a high performance liquid chromatograph with fluorescence detector. The chromatographic conditions were as follows: an ODS column (4.6 mm i.d. x 150 mm, 5 microns), was eluted with a flow rate of 1.0 mL/min. The fluorescence detector was operated at lambda ex 365 nm and lambda em 475 nm. Mobile phase frompump A was 3% methanol containing 0.25% acetic acid. In gradient elution program the methanol from pump B was as follows: 0-8 min, 5%; 8-15 min, 5%-12%; 15-20 min, 12%; 20-30 min, 12%-20%; 30-32 min, 20%; 32-35 min, 20%-100%; 35-40 min, 100%; 40-43 min, 100%-5%; 43-45 min, 5%. The method proved to be linear in the range of 2.5-80.0 mumol/L with a regression coefficient of 0.9988. The minimum detection limit was 0.5 mumol/L, the recoveries were 94.0%-112.0%, and the RSD values were less than 5.6%.

Chromatography, High Pressure Liquid↗
PubMed1976

Analysis of variation in automated determination of sodium, potassium and calcium ions in human serum.

The concentration of sodium ion (Na+), potassium ion (K+) and calcium ion (Ca2+) in patient sera were assayed simultaneously on two separate flame photometers and then evaluated by analysis of variance (ANOVA). Two different experimental designs were devised in order to separate the error components assignable to pipetting, dilution, and instrumental measurement. Within-series variability was compared with the between-series variability. Similar statistical treatment of results from quality control samples gave estimates of variances between days, between dilutions, between instruments, and between channels, as well as the interaction terms and instrumental measurement errors. The total (overall) coefficients of variation (CV) were 1.15, 1.60, and 1.38% for Na+, K+, and Ca2+, respectively. The adequacy of the quality-control system to predict expected variation in patient specimens was verified by comparing errors from patient-sample duplicates and from control samples.

Analysis of Variance↗
PubMed1992

[Emoxipin as an inhibitor of angiogenesis].

The effect of emoxypin on angiogenesis in rabbit cornea in aseptic inflammation induced by intracorneal implantation of a piece of quartz and on the development of the vessels of the chick embryo yolk sac was studied. 1% emoxypin pipetted thrice a day for 10-14 days inhibited corneal neovascularization and reduced the formation of new blood vessels. We observed an inhibitory effect on the development of vascular bed of the embryo yolk sac on incubation hour 64-72. The drug affected neither general growth of the embryos no the number of somites.

Animals↗
PubMed1992

[Electronic data processing-assisted serial automation of current methods in blood group serology].

The introduction of special centrifugable racks with a transparent bottom into the conventional typing of blood groups in glass tubes facilitates the simultaneous work on and reading of a maximum of 32 complete ABO, Rhesus and Kell typings in one series. As a result of the facts that it is unnecessary to label the individual tubes and that the pipetting of serum and erythrocyte suspensions is done automatically and through the unmistakable classification of the samples by means of bar-coding, the manual work is reduced to about 50%. This modified typing method combines the advantages of the conventional typing method in glass tubes with the advantages of an efficient microplate technique. There is no qualitative loss in the implementation and reading of the analysis.

ABO Blood-Group System↗
PubMed2003

Description and classification of abnormal cercariae of Schistosoma mattheei and a method of concentration and collection.

Over a period of about 12 years, 30 abnormal Schistosoma mattheei cercariae were found among a total of approximately 2.8 million examined. Initially seven were recovered from about 1.02 million (0.0007%), which were examined individually while being counted with the aid of a stereoscopic microscope. Subsequently, on the strength of relatively high percentages of abnormal individuals recovered when counting cercariae that failed to penetrate into oxen, it appeared that the morphologically abnormal cercariae were unable to swim and would mostly sediment out of a suspension while most of the normal cercariae would remain swimming. This surmise is supported by recovery of 23 morphologically abnormal cercariae (0.001%) from about 1.8 million, by examining the sediment after the cercarial suspension had been left standing undisturbed in glass measuring cylinders. The abnormalities ranged from aberrant tails only (e.g. an underdeveloped tail, or different degrees of schism) or aberrant heads only, to abnormalities of both the heads and tails. A suggested schematic classification of abnormal cercariae is presented. A young, adult hamster was exposed to eight S. mattheei cercariae with complete schism of the shaft of the tail, by pipetting the cercariae onto the shaved abdominal skin of the anaesthetised animal. Two underdeveloped females were subsequently encountered in squash preparations of the liver when the hamster was killed for worm recovery 10 weeks after infection, thus showing that some of the abnormal cercariae were viable. A method is also described for killing and fixing cercariae while retaining some of the shining brilliance of live cercariae, without them becoming shrivelled, granular and semi-opaque, as occurs when cercariae die spontaneously or are killed with heat. This is apparently the first report of abnormal cercariae of S. mattheei. In addition, a method of concentrating abnormal cercariae after emergence from a snail, a schematic classification of abnormal cercariae and a method for killing and fixing cercariae while retaining much of the shiny brilliance of live cercariae are also reported for the first time as far as is known.

Animals↗
PubMedDate not supplied

Simultaneous quantitation of cytokine mRNAs in interleukin-1 beta stimulated U373 human astrocytoma cells by a polymerisation chain reaction method involving co-amplification with an internal multi-specific control.

The measurement of cytokine mRNA levels is of fundamental importance in the understanding of diverse pathological states. We present a simplification of a polymerase chain reaction-based technique which permits the simultaneous measurement of up to 20 cytokine mRNAs, together with those of several other cellular products, including beta 2-microglobulin and beta-actin. The technique makes use of internal standards bearing multiple PCR primer sites which are identical to those on the mRNAs to be assayed. Known quantities of the standards are added to the cellular RNA and the mixture is co-reverse transcribed and co-amplified. The simplifications described here are based on the fact that each pair of amplicons accumulates in a constant ratio even in the plateau phase of amplification. As a result, no preliminary experiments to determine the limits of the exponential phase of amplification are necessary; the same number of cycles may be chosen for all the mRNAs to be measured, whatever their level in the mixture might be; pipetting errors are avoided since all calculations are based upon the relative quantities of co-amplified material. Here we illustrate the method through a quantitative study of the expression of cytokine mRNAs in U373 human astrocytoma cells before and after stimulation with IL-1 beta. Quantitation was carried out either by incorporating radioactivity in the amplicons or by fluorescence measurements after propidium iodide staining. Only very low numbers of transcripts for IL-6, IL-8, CSF-1, MCP-1 and either Gro alpha or Gro beta were detectable in unstimulated cells. The levels of these cytokine mRNAs increased dramatically following IL-1 beta stimulation and, in addition, transcription of IL-1 beta, TNF alpha, GM-CSF, G-CSF, Gro gamma and MCP-1, some of which have not previously been detected in U373, was initiated in the stimulated cells. At the same time we found that transcripts for IL-2, IL-3, IL-4, IL-5, IFN gamma, huMlP1 alpha and huMlP1 beta were totally absent in this cell line. These results suggest a potentially important role for astrocytes in the local amplification of inflammatory responses in the brain.

Astrocytes↗
PubMedDate not supplied

[B viral hepatitis as a risk factor for clinical laboratory personnel].

A study was carried out on the risk of contamination with viral hepatitis B for the personnel of the clinical laboratories who handle blood samples with HB Ag and HB Ac, collected from the patients. Of the total 1945 blood samples collected from 639 adult patients, 74 were serologically positive (3.8%) of which 39 HB Ag carriers (2%) and 35 HB Ac carriers (1.8%). Although these proportions are close to those found in the normal population the risk of contamination of the laboratory personnel is greater owing to the direct contact with the pathologic product and the impossibility of recognizing the HB Ag and Ac positive samples before testing. The risk may be lowered by strict compliance with the general rules of hygiene, by avoiding especially any microtraumas of the hands, by automatic pipetting of the sera and reagents and the use of sterilized materials (particularly in haematologic and biochemical laboratories.

Hepatitis A↗